Anti-NSCLC activity in vitro of Hsp90^N inhibitor KW-2478 and complex crystal structure determination of Hsp90^N-KW-2478

KW-2478 is a promising anti-cancer lead compound targeting to the molecular chaperone heat shock protein 90(N) (Hsp90(N)). Absence of complex crystal structure of Hsp90(N)-KW-2478, however, hampered further structure optimization of KW-2478 and understanding on the molecular interaction mechanism. Herein, a high-resolution complex crystal structure of Hsp90(N)-KW-2478 was determined by X-ray diffraction (XRD, resolution limit: 1.59 angstrom; PDB ID: 6LT8) and their molecular interaction was analyzed in detail, which suggested that KW-2478 perfectly bound in the N-terminal ATP-binding pocket of Hsp90 to disable its molecular chaperone function, therefore suppressed or killed cancer cells. The results from thermal shift assay (TSA, Delta Tm, 18.82 +/- 0.51 degrees C) and isothermal titration calorimetry (ITC, K-d, 7.30 +/- 2.20 nM) suggested that there is an intense binding force and favorable thermodynamic changes during the process of KW-2478 binding with Hsp90(N). Additionally, KW-2478 exhibited favorable anti-NSCLC activity in vitro, as it inhibited cell proliferation (IC50, 8.16 mu M for A549; 14.29 mu M for H1975) and migration, induced cell cycle arrest and promoted apoptosis. Thirty-six novel KW-2478 derivatives were designed, based on the complex crystal structure and molecular interaction analysis of Hsp90(N)-KW-2478 complex. Among them, twenty-two derivatives exhibited increased binding force with Hsp90(N) evaluated by molecular docking assay. The results would provide new guidance for anti-NSCLC new drug development based on the lead compound KW-2478.